A previously uncharacterized Factor Associated with Metabolism and Energy (FAME/C14orf105/CCDC198/1700011H14Rik) is related to evolutionary adaptation, energy balance, and kidney physiology

In this study we use comparative genomics to uncover a gene with uncharacterized function (1700011H14Rik/C14orf105/CCDC198), which we hereby name FAME (Factor Associated with Metabolism and Energy). We observe that FAME shows an unusually high evolutionary divergence in birds and mammals. Through the comparison of single nucleotide polymorphisms, we identify gene flow of FAME from Neandertals into modern humans. We conduct knockout experiments on animals and observe altered body weight and decreased energy expenditure in Fame knockout animals, corresponding to genome-wide association studies linking FAME with higher body mass index in humans. Gene expression and subcellular localization analyses reveal that FAME is a membrane-bound protein enriched in the kidneys. Although the gene knockout results in structurally normal kidneys, we detect higher albumin in urine and lowered ferritin in the blood. Through experimental validation, we confirm interactions between FAME and ferritin and show co-localization in vesicular and plasma membranes.


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March 2021

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Sample size -Library preparation was performed using a 10x controller (10x Genomics) with the Single Cell 3' v3 chemistry. Sequencing was performed using a HiSeq 3000 (Illumina) -LC-MS/MS analyses of peptide mixture were done using Ultimate 3000 RSLCnano system connected to Orbitrap Elite hybrid spectrometer (Thermo Fisher Scientific).
-For 3D visualization, the segmentation was done by an operator using a combination of software Avizo 2022.2 (ThermoFisher Scientific) and VG Studio MAX 3.4 (Volume Graphics GmbH, Germany).
Statistical data analysis: Statistical analysis was done using GraphPad Prism 9 software.
The quantitative data are provided in the Source Data file.
All custom-made scripts used in the analysis are available at: https://github.com/ipoverennaya/RIK_paper Two knockout mice strains were generated for this manuscript. These will be available upon reasonable request and also be deposited to the Jackson Laboratory. All other relevant data supporting the key findings of this study are available within the article and its Supplementary Information files or from the corresponding author upon reasonable request. The quantitative data generated in this study are provided in the Source Data file. Source data are provided with this paper. n/a n/a n/a n/a In our study, we did not perform a formal sample size calculation. Instead, we followed accepted practices in the mouse facilities where the research was conducted to determine the sample size. We initially conducted preliminary experiments to estimate the number of animals needed to achieve adequate statistical power for our tests. Furthermore, the number of animals involved in this research were chosen to reduce unnecessary animal use and to reach statistical significance with two-sided student´s t-test. For the mass spectrometry experiments, we analyzed six independent replicates to ensure the reliability and reproducibility of our findings. For immunofluorescence, single cell sequencing, and transmission electron microscopy, we used three independent samples each. These sample sizes were selected based on the Validation same approach of balancing statistical power and accepted practices in the research community to yield robust effects.
To check whether DE genes between wildtype and knockout are not sex-specific, we compared them with the list of the corresponding DE genes between female and male proximal tubule (PT) samples from (Ransick et al., 2019). The genes whose adjusted p-values are less than 0.01 were excluded from the comparative analysis For plotted graphs we did not exclude any data.
In vitro experiments were repeated a minimum of three times. All attempts were succesful.
Parameters related to mice were tested once in several animals (minimum of four). All attempts were succesful.
Laboratory animals were allocated to the experimental and control groups based on their genotype and sex. The same accounted for the tissues harvested from these mice.
In vitro experiments were allocated based on their treatment (control vs. inhibitor) or transfection condition.
Blinding was not applicable to the study because the allocation of laboratory animals to experimental and control groups was based on their genotype and sex, and the tissues harvested from these animals were also allocated accordingly. Additionally, the in vitro experiments were allocated based on treatment (control vs. inhibitor) or transfection condition. Lotus tetragonolobus lectin: Lotus tetragonolobus lectin (LTL) encompasses a family of closely related glycoproteins with similar specificities toward "-linked L-fucose-containing oligosaccharides. Although many of the binding properties of Lotus lectin are similar to those of Ulex europaeus lectin I (UEL I), the binding affinities and some specificities for oligosaccharides are significantly different between these fucose-specific lectins. This fluorescein-labeled LTL features a ratio of fluorophores to lectin protein that provides optimal staining (excitation 495 nm, emission 515 nm). Supplied as a solution essentially free of unconjugated fluorophores, it is preserved with sodium azide. The recommended inhibiting/eluting sugar is 50-100 mM L-fucose. https://doi.org/10.1016/j.celrep.2022.110473 Anti-VANGL1: Vangl1 Antibody is a mouse monoclonal IgG1 # Vangl1 antibody provided at 200 µg/ml specific for an epitope mapping between amino acids 281-298 within a cytoplasmic domain of Vangl1 of human origin. Vangl1 Antibody is recommended for detection of Vangl1 of mouse, rat and human origin by WB, IP, IF and ELISA; also reactive with additional species, including and equine, canine, bovine and porcine https://doi.org/10.1371/journal.pgen.1007840 anti-GFP: Chicken polyclonal antibody to GFP with over 2500 references: https://www.abcam.com/products/primary-antibodies/gfpantibody-ab13970.html?productWallTab=ShowAll